Dividing cells undergo a cyclical programmed that culminates in cell division. A normal event in this cell cycle programmed is the replication of the cellular DNA. Just prior to DNA synthesis, there is a pause to allow proof reading of the DNA, ensuring that any damage or mutations are repaired and not passed on to daughter cells. This checkpoint (GI) is regulated by programmed gene expression. A second, similar checkpoint occurs later on after DNA synthesis, just before the cell divides into two new cells, presumably for the same purpose. Senescent or aged cells are permanently arrested at one of these checkpoints. Recently, Olivia Pereira-Smith, Jim Smith and colleagues have identified three cDNA's that cause growth arrest when transfected into young, actively dividing cells (Noda, A., et al., Exp. Cell. Res., 211:900–98 (1994)). Other groups have described SDI-1 as WAF1, CIP1; PIC1 and p21 (Harper, J. N., et al., Cell, 75:805–816 (1993); El-Deiry, W. S. et al., Cell, 75:817–825 (1993); Xiong, Y., et al., Nature, 366:701–704 (1993); Hunter, T. et al., Cell, 75:839–841 (1993)). This gene plays a central role in cellular processes that have in common the loss of cell proliferation which implicates this gene as being involved in cell cycle control. SDI-1 has been shown to be overexpressed in senescent cells, quiescent cells or cultured primary cells undergoing crisis (Noda, A., et al., Exp. Cell Res., 211:900–98 (1994); Rubelj, I. and Pereira-Smith, O. M., Exp. Cell Res. 211:82–89 (1994)), suggesting a role in the maintenance of DNA synthesis inhibition (Johnson, M., et al., Mol. Carcinogen., 11:59–64 (1994)). Further, evidence has been presented suggesting that the SDI-1 medicated inhibition of DNA synthesis occurs via an inhibition of CDK (cyclin-dependent kinases) activity (Nakanishi, M., et al., Proc. Natl. Acad. Sci. USA, 92:4352–4356 (1995)). These findings, together with the demonstration that SDI-1 can inhibit cell growth of young dividing cells, suggests that this gene will be useful for gene therapy to inhibit the growth of rapidly proliferating cells in diseases or disorders such as restenosis, in which smooth muscle cells inappropriately divide, or various cancers.
In this respect, SDI-1 will be useful therapeutically to suppress the rapid proliferation of tumor or tumorigenic cells, such as breast, lung, hepatic and glioma tumor cells. SDI-1 has also been described to have the ability to mediate the differentiation of cancer cells (malignant melanoma cells) into non-cancerous cells. Additional applications for SDI-1 in a therapeutic context include the inhibition of endothelial cell replication (antiangiogenesis) to prevent neovascularizations of tumors, and to halt cell growth (WO-Al-95 06415).
SDI-1 induces a senescent or quiescent state in recipient cells and will therefore also be useful in the treatment of various age-related disorders, asthenia and cachexia, or other diseases or conditions in which rapid cellular proliferation is undesirable.
SDI-1 can also be used in combination with conventional chemotherapeutic agents in order to enhance the efficacy of chemotherapeutic agents. The premise of chemotherapy is that cancer cells grow more rapidly than normal cells, and hence are more sensitive to cytotoxic agents than normal cells. Many chemotherapeutic agents exert their effect during a specific phase or set of phases of the cell cycle, and because only a fraction of tumor cells are in a specific phase at any given time, such drugs must generally be provided by repeated administration.
SDI-1 can be used to synchronize or maximize the percentage of cells that are in a particular phase of the cell cycle at the time of administering the chemotherapeutic agent, and thereby provide a means to increase the effectivity of chemotherapy.
Other diseases such as psoriasis, rheumatoid arthritis and restenosis can also be treated with SDI-1. The use of SDI-1 for the treatment of viral and microbial infections and several other disorders or diseases are described in WO-Al-95/06415.
Antisense sequences are nucleic acids (either DNA or RNA) whose sequence is complementary to the sequence of a target mRNA molecule (or its corresponding gene) so that it is capable of hybridizing with binding to the mRNA molecule (or gene) and thereby impairing (i.e. attenuating or preventing) the transcription of the gene into mRNA or the translation of the mRNA molecule into a gene product.
Antisense SDI-1 DNA sequences may thus be used to inhibit the production of endogenous SDI-1 and thereby stimulate the proliferation of cells. Antisense SDI-1 DNA sequences may therefore be used to promote wound healing, angiogenesis, endothelial cell proliferation, recovery from burns or surgery or to restore attrophied tissue (WO-Al-95 06415). Antisense SDI-1 DNA sequences can also be used to immortalize cells and permit the establishment of permanent cell lines. Antisense SDI-1 DNA sequences may also be used in research or to permit or facilitate the accumulation of large numbers of cells, as for organ tissue grafts or transplants, or they may be used to immortalize cells producing important biological molecules such as hormones, interferons, or growth factors.
Transcription of antisense SDI-1 DNA sequences in cells in combination with irradiation leads to an increase in cell death. Transcription of antisense SDI-1 DNA sequences may therefore also be useful for gene therapy for the treatment of diseases or disorders characterized by rapid cell proliferation, such as certain tumors as well as in vascular diseases such as restenosis.
Retroviral vectors are suitable gene transfer vehicles for stable therapeutic gene delivery to rapidly dividing cells such as cancer cells, becuase most retroviral particles only infect dividing cells and not the surrounding non-dividing cells. Further, the retroviral vector integrates in the chromosomal DNA of dividing cells, thereby ensuring the transmission of therapeutic genes to all daughter cells, if the original infected cell is proliferating. Retroviral vectors are therefore the ideal vector for the delivery of DNA sequences encoding polypeptides affecting the proliferation of cells, such as SDI-1, into cells.